392 research outputs found

    LAB-1 Targets PP1 and Restricts Aurora B Kinase upon Entrance into Meiosis to Promote Sister Chromatid Cohesion

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    Successful execution of the meiotic program depends on the timely establishment and removal of sister chromatid cohesion. LAB-1 has been proposed to act in the latter by preventing the premature removal of the meiosis-specific cohesin REC-8 at metaphase I in C. elegans, yet the mechanism and scope of LAB-1 function remained unknown. Here we identify an unexpected earlier role for LAB-1 in promoting the establishment of sister chromatid cohesion in prophase I. LAB-1 and REC-8 are both required for the chromosomal association of the cohesin complex subunit SMC-3. Depletion of lab-1 results in partial loss of sister chromatid cohesion in rec-8 and coh-4 coh-3 mutants and further enhanced chromatid dissociation in worms where all three kleisins are mutated. Moreover, lab-1 depletion results in increased Aurora B kinase (AIR-2) signals in early prophase I nuclei, coupled with a parallel decrease in signals for the PP1 homolog, GSP-2. Finally, LAB-1 directly interacts with GSP-1 and GSP-2. We propose that LAB-1 targets the PP1 homologs to the chromatin at the onset of meiosis I, thereby antagonizing AIR-2 and cooperating with the cohesin complex to promote sister chromatid association and normal progression of the meiotic program

    Direct observation of mammalian cell growth and size regulation

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    We introduce a microfluidic system for simultaneously measuring single cell mass and cell cycle progression over multiple generations. We use this system to obtain over 1,000 hours of growth data from mouse lymphoblast and pro-B-cell lymphoid cell lines. Cell lineage analysis revealed a decrease in the growth rate variability at the G1/S phase transition, which suggests the presence of a growth rate threshold for maintaining size homeostasis

    Automating a framework to extract and analyse transport related social media content: The potential and the challenges

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    Harnessing the potential of new generation transport data and increasing public participation are high on the agenda for transport stakeholders and the broader community. The initial phase in the program of research reported here proposed a framework for mining transport-related information from social media, demonstrated and evaluated it using transport-related tweets associated with three football matches as case studies. The goal of this paper is to extend and complement the previous published studies. It reports an extended analysis of the research results, highlighting and elaborating the challenges that need to be addressed before a large-scale application of the framework can take place. The focus is specifically on the automatic harvesting of relevant, valuable information from Twitter. The results from automatically mining transport related messages in two scenarios are presented i.e. with a small-scale labelled dataset and with a large-scale dataset of 3.7 m tweets. Tweets authored by individuals that mention a need for transport, express an opinion about transport services or report an event, with respect to different transport modes, were mined. The challenges faced in automatically analysing Twitter messages, written in Twitter’s specific language, are illustrated. The results presented show a strong degree of success in the identification of transport related tweets, with similar success in identifying tweets that expressed an opinion about transport services. The identification of tweets that expressed a need for transport services or reported an event was more challenging, a finding mirrored during the human based message annotation process. Overall, the results demonstrate the potential of automatic extraction of valuable information from tweets while pointing to areas where challenges were encountered and additional research is needed. The impact of a successful solution to these challenges (thereby creating efficient harvesting systems) would be to enable travellers to participate more effectively in the improvement of transport services

    Rapid generation of endogenously driven transcriptional reporters in cells through CRISPR/Cas9

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    CRISPR/Cas9 technologies have been employed for genome editing to achieve gene knockouts and knock-ins in somatic cells. Similarly, certain endogenous genes have been tagged with fluorescent proteins. Often, the detection of tagged proteins requires high expression and sophisticated tools such as confocal microscopy and flow cytometry. Therefore, a simple, sensitive and robust transcriptional reporter system driven by endogenous promoter for studies into transcriptional regulation is desirable. We report a CRISPR/Cas9-based methodology for rapidly integrating a firefly luciferase gene in somatic cells under the control of endogenous promoter, using the TGFβ-responsive gene PAI-1. Our strategy employed a polycistronic cassette containing a non-fused GFP protein to ensure the detection of transgene delivery and rapid isolation of positive clones. We demonstrate that firefly luciferase cDNA can be efficiently delivered downstream of the promoter of the TGFβ-responsive gene PAI-1. Using chemical and genetic regulators of TGFβ signalling, we show that it mimics the transcriptional regulation of endogenous PAI-1 expression. Our unique approach has the potential to expedite studies on transcription of any gene in the context of its native chromatin landscape in somatic cells, allowing for robust high-throughput chemical and genetic screens

    A Stepwise Analytical Projected Gradient Descent Search for Hyperspectral Unmixing and Its Code Vectorization

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    We present, in this paper, a new methodology for spectral unmixing, where a vector of fractions, corresponding to a set of endmembers (EMs), is estimated for each pixel in the image. The process first provides an initial estimate of the fraction vector, followed by an iterative procedure that converges to an optimal solution. Specifically, projected gradient descent (PGD) optimization is applied to (a variant of) the spectral angle mapper objective function, so as to significantly reduce the estimation error due to amplitude (i.e., magnitude) variations in EM spectra, caused by the illumination change effect. To improve the computational efficiency of our method over a commonly used gradient descent technique, we have analytically derived the objective function's gradient and the optimal step size (used in each iteration). To gain further improvement, we have implemented our unmixing module via code vectorization, where the entire process is ''folded'' into a single loop, and the fractions for all of the pixels are solved simultaneously. We call this new parallel scheme vectorized code PGD unmixing (VPGDU). VPGDU has the advantage of solving (simultaneously) an independent optimization problem per image pixel, exactly as other pixelwise algorithms, but significantly faster. Its performance was compared with the commonly used fully constrained least squares unmixing (FCLSU), the generalized bilinear model (GBM) method for hyperspectral unmixng, and the fast state-of-the-art methods, sparse unmixing by variable splitting and augmented Lagrangian (SUnSAL) and collaborative SUnSAL (CLSUnSAL) based on the alternating direction method of multipliers. Considering all of the prospective EMs of a scene at each pixel (i.e., without a priori knowledge which/how many EMs are actually present in a given pixel), we demonstrate that the accuracy due to VPGDU is considerably higher than that obtained by FCLSU, GBM, SUnSAL, and CLSUnSAL under varying illumination, and is, otherwise, comparable with respect to these methods. However, while our method is significantly faster than FCLSU and GBM, it is slower than SUnSAL and CLSUnSAL by roughly an order of magnitude.Israel Science Ministry Scientific Infrastructure Research Grant Scheme, Helen Norman Asher Space Research Grant Scheme, Technion PhD Scholarship, new England fund Technion, Environmental Mapping and Monitoring of Iceland by Remote Sensing EMMIRS projectPeer Reviewe

    Optimizing Optical Flow Cytometry for Cell Volume-Based Sorting and Analysis

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    Cell size is a defining characteristic central to cell function and ultimately to tissue architecture. The ability to sort cell subpopulations of different sizes would facilitate investigation at genomic and proteomic levels of mechanisms by which cells attain and maintain their size. Currently available cell sorters, however, cannot directly measure cell volume electronically, and it would therefore be desirable to know which of the optical measurements that can be made in such instruments provide the best estimate of volume. We investigated several different light scattering and fluorescence measurements in several different cell lines, sorting cell fractions from the high and low end of distributions, and measuring volume electronically to determine which sorting strategy yielded the best separated volume distributions. Since we found that different optical measurements were optimal for different cell lines, we suggest that following this procedure will enable other investigators to optimize their own cell sorters for volume-based separation of the cell types with which they work

    The Genographic Project Public Participation Mitochondrial DNA Database

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    The Genographic Project is studying the genetic signatures of ancient human migrations and creating an open-source research database. It allows members of the public to participate in a real-time anthropological genetics study by submitting personal samples for analysis and donating the genetic results to the database. We report our experience from the first 18 months of public participation in the Genographic Project, during which we have created the largest standardized human mitochondrial DNA (mtDNA) database ever collected, comprising 78,590 genotypes. Here, we detail our genotyping and quality assurance protocols including direct sequencing of the mtDNA HVS-I, genotyping of 22 coding-region SNPs, and a series of computational quality checks based on phylogenetic principles. This database is very informative with respect to mtDNA phylogeny and mutational dynamics, and its size allows us to develop a nearest neighbor–based methodology for mtDNA haplogroup prediction based on HVS-I motifs that is superior to classic rule-based approaches. We make available to the scientific community and general public two new resources: a periodically updated database comprising all data donated by participants, and the nearest neighbor haplogroup prediction tool
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